Benzylpiperazine: "A messy drug".

Designer drugs are synthetic structural analogues/congeners of controlled substances with slightly modified chemical structures intended to mimic the pharmacological effects of known drugs of abuse so as to evade drug classification. Benzylpiperazine (BZP), a piperazine derivative, elevates synaptic dopamine and serotonin levels producing stimulatory and hallucinogenic effects, respectively, similar to the well-known drug of abuse, methylenedioxymethamphetamine (MDMA). Furthermore, BZP augments the release of norepinephrine by inhibiting presynaptic autoreceptors, therefore, BZP is a "messy drug" due to its multifaceted regulation of synaptic monoamine neurotransmitters. Initially, pharmaceutical companies used BZP as a therapeutic drug for the treatment of various disease states, but due to its contraindications and abuse potential it was withdrawn from the market. BZP imparts predominately sympathomimetic effects accompanied by serious cardiovascular implications. Addictive properties of BZP include behavioral sensitization, cross sensitization, conditioned place preference and repeated self-administration. Additional testing of piperazine derived drugs is needed due to a scarcity of toxicological data and widely abuse worldwide.

Novel inhibitors of rat lens aldose reductase: N-[[(substituted amino)phenyl]sulfonyl]glycines.

A number of N-[[(substituted amino)phenyl]sulfonyl]glycines 3a-n were synthesized as analogues of the simple (phenylsulfonyl)glycines 1a-c with increased lipophilic character and therefore greater aldose reductase inhibitory potential. The 2-benzoylamino derivative 3c was found to be less potent than the corresponding amine 1c as an inhibitor of rat lens aldose reductase, but both the 3- and 4-benzoylamino analogues, 3b and 3a, are substantially more potent than their amines 1b and 1a; compound 3a is the most effective inhibitor of this series, with an IC50 of 0.41 microM. The 4-benzoylamino derivative 3a is also significantly more active than the 4-acetylamino analogue 3d and the 4-benzylamino (3e) and 4-dimethylamino (3f) derivatives, suggesting that both the additional carbonyl moiety and aromatic ring present in this compound may bind to complementary sites present on the enzyme. Furthermore, structure-activity studies reveal that increasing the number of atoms between the carbonyl and aromatic moieties of 3a results in a decrease in inhibitory activity. Kinetic studies demonstrate that 3a, like other known inhibitors of aldose reductase, functions as an uncompetitive inhibitor with respect to the substrate and therefore may interact at the proposed common inhibitor binding site of this enzyme.

N- and 2-substituted N-(phenylsulfonyl)glycines as inhibitors of rat lens aldose reductase.

A variety of N-(phenylsulfonyl)-N-phenylglycines 5, N-(phenylsulfonyl)-2-phenylglycines 6, and N-(phenylsulfonyl)anthranilic acids 7 were prepared as analogues of the N-(phenylsulfonyl)glycine 1 aldose reductase inhibitors. In the rat lens assay, several derivatives of 5 display greater inhibitory activity than the corresponding glycines 1, suggesting that N-phenyl substitution enhances affinity for aldose reductase. Enzyme kinetic evaluations of the 4-benzoylamino analogues of 5 and 1 demonstrate that these compounds produce inhibition by the same mechanism. However, the significant differences in relative inhibitory potencies between compounds of series 5 and 1 may indicate that these compounds do not interact with the inhibitor binding site in precisely the same manner. Evaluation of the individual enantiomers of series 6 reveals that the S isomers are substantially more active than the corresponding R isomers. Also, with the exception of the naphthalene analogue 6n, the S stereoisomers of this series display greater inhibitory potencies than the glycines 1. The anthranilates 7 generally are less active than the glycines 1, demonstrating that direct incorporation of an aromatic ring in the glycine side chain may result in a decrease in affinity for aldose reductase.

Synthesis and rat lens aldose reductase inhibitory activity of some benzopyran-2-ones.

A number of 4,7-disubstituted benzopyran-2-ones were synthesized and evaluated for crude rat lens aldose reductase inhibitory activity. Substituents on position 4 included CH3, CO2H, CH2CO2H, CH = CHCO2H, and CH2CH2CO2H. The aromatic substituents included OH, OCH3, OCOCH3, CH2CH3, and Cl. Also included in the study were 3-oxo-3H-naphtho[2,1-b]pyran-1-acetic, 2-oxo-2H-naphtho[1,2-b]pyran-4-acetic, and 1-naphthylacetic acids. The benzopyran and naphthopyran derivatives were prepared by the classical von Pechmann reaction. General structure-activity relationships reveal that optimal enzyme inhibitory activity is displayed by those compounds possessing the acetic acid moiety. For example, the most potent derivative, 3-oxo-3H-naphtho[2,1-b]pyran-1-acetic acid with an IC50 of 0.020 microM, is as potent as sorbinil (IC50 = 0.017 microM) in the crude rat lens aldose reductase assay.

Synthesis and in vitro aldose reductase inhibitory activity of compounds containing an N-acylglycine moiety.

A number of N-benzoylglycines (6), N-acetyl-N-phenylglycines (7), N-benzoyl-N-phenylglycines (8), and tricyclic N-acetic acids (9-12) were synthesized as analogues of the N-acylglycine-containing aldose reductase inhibitors alrestatin and 2-oxoquinoline-1-acetic acid. Derivatives of 6, which represent ring-simplified analogues of alrestatin, are very weak inhibitors of aldose reductase obtained from rat lens, producing 50% inhibition only at concentrations exceeding 100 microM. Compounds of series 7 were designed as ring-opened analogues of the 2-oxoquinolines. While these derivatives are more potent than compounds of series 6 (IC50S of 6-80 microM), they are less active than the corresponding 2-oxoquinolines. Analogues of series 8 were designed as hybrid structures of both alrestatin and the 2-oxoquinoline-1-acetic acids. These compounds are substantially more potent than compounds of series 6 and 7 and display inhibitory activities comparable to or greater than alrestatin or the 2-oxoquinolines (IC50S of 0.1-10 microM). Of the rigid analogues of 8, the most potent derivative is benzoxindole (12) with an IC50 of 0.67 microM, suggesting that fusion of the two aromatic rings of 8 in a coplanar conformation may optimize affinity for aldose reductase in this series.

Relative structure-inhibition analyses of the N-benzoyl and N-(phenylsulfonyl) amino acid aldose reductase inhibitors.

A number of N-benzoyl amino acids were synthesized and tested to compare structure-inhibition relationships with the isosteric N-(phenylsulfonyl) amino acid (PS-amino acid) aldose reductase inhibitors. Inhibition analyses with these series reveals that their kinetic mechanisms of inhibition are similar, but that significant differences in structure-inhibition relationships exist. For example, while the PS-alanines and PS-2-phenylglycines produce enantioselective inhibition (S greater than R), no consistent pattern of enantioselectivity is observed with the isosteric N-benzoylalanines and 2-phenylglycines. Also, N-methyl and N-phenyl substitution in the PS-amino acid series does not substantially alter inhibitory activity, while similar substitutions in the N-benzoyl series (particularly N-phenyl) results in a significant increase in inhibitory activity. Proton NMR analysis of the N-benzoylsarcosines reveals that these compounds exist as a mixture of rotamers in solutions including the enzyme assay buffer and that the preferred conformer is one in which the carboxymethyl moiety is trans to the aromatic ring. Similar analyses with the N-benzoyl-N-phenylglycines demonstrate that these derivatives exist exclusively in the trans rotameric conformation in solution. No such N-substituent effects on conformation were observed in the PS-amino acid series. These results suggest that the differences in structure-inhibition trends between these structurally related series may result from the effect of substituents on preferred conformation.

Design and synthesis of 2-(arylamino)-4(3H)-quinazolinones as novel inhibitors of rat lens aldose reductase.

A number of 2-(arylamino)-4(3H)-quinazolinones (2a-i) that possess several of the pharmacophore moieties necessary for binding to the inhibitor site of the enzyme aldose reductase were synthesized and tested for their ability to inhibit crude aldose reductase obtained from rat lens. Only those quinazolinones that possess an acidic moiety on the 2-(arylamino) substituent were found to display significant inhibitory activity. Of these, the most potent compound is the 4'-CO2H derivative (2i) with an IC50 of 34 microM, while the least potent is the 4'-OH derivative (2c) with an IC50 of 75 microM. All of the 2-(arylamino)-4(3H)-quinazolinones tested are less potent than other known inhibitors of aldose reductase, such as alrestatin and sorbinil, indicating that the pharmacophore moieties present in these compounds may not be positioned optimally relative to one another for maximal interaction with the enzyme.

Synthesis and aldose reductase inhibitory activity of substituted 2-oxoquinoline-1-acetic acid derivatives.

A number of 2-oxoquinoline-1-alkanoic acids that contain the N-acylglycine fragment found in several known inhibitors of aldose reductase were synthesized and tested in the rat lens assay. All of the target compounds were prepared by alkylation of the appropriate 2-oxoquinoline intermediates with a halo ester, followed by hydrolysis of the intermediate esters. In the rat lens assay, the 1-acetic acid derivatives 9a-e display the highest level of aldose reductase inhibitor activity with IC50 values of 0.45-6.0 microM. Modification of the 1-acetic acid moiety by esterification, substitution of an alpha-methyl group, or insertion of an additional methylene unit results in reduced inhibitory potency. Structure-activity data also suggests that both the benzene and 2-oxopyridine rings of 9a-e contribute substantially toward activity and that inhibitory potency is influenced by aromatic ring substituents.

Inhibitory activity and mechanism of inhibition of the N-[[(4-benzoylamino)phenyl]sulfonyl]amino acid aldose reductase inhibitors.

A series of substituted N-[[(4-benzoylamino)phenyl]sulfonyl]amino acids (BAPS-amino acids) were synthesized by established methods, and the stereochemistry of the products was confirmed by HPLC analysis after chiral derivatization. When tested against aldose reductase (alditol:NADP+ oxidoreductase; EC 1.1.1.21; ALR2) isolated from rat lens, all of the BAPS-amino acids were determined to be significantly more inhibitory than the corresponding N-(phenylsulfonyl)amino acids. Structure-inhibition and enzyme kinetic analyses suggest that the BAPS-amino acids inhibit ALR2 by a mechanism similar to the N-(phenylsulfonyl)amino acids. However, multiple inhibition analyses indicate that the increased inhibitory activity of the BAPS-amino acids is a result of interaction with multiple sites present on ALR2. Enzyme specificity studies with several of the BAPS-amino acids demonstrated that these compounds do not produce significant inhibition of other nucleotide-requiring enzymes including aldehyde reductase (alcohol: NADP+ oxidoreductase; EC 1.1.1.2; ALR1).

Behavioral and developmental effects of two 3,4-methylenedioxymethamphetamine (MDMA) derivatives.

The effects of 3,4-methylenedioxymethamphetamine (MDMA or 'ecstacy') and two structurally related compounds, N-methyl-1-(3,4-methylenedioxyphenyl)-1-ethanamine (MDM1EA) and N-methyl-1-(3,4-methylenedioxyphenyl)-3-butanamine (HMDMA) were examined in two preparations: (i) a drug discrimination procedure in MDMA-trained rats and (ii) the chicken embryo, for determination of the direct effects of these compounds on the developing organism. The highest doses of MDM1EA and HMDMA partially substituted for MDMA, whereas higher (30-60 mg/kg) doses of HMDMA evoked clonic seizures in a separate group of rats. In chicken embryos MDMA had no effect on body, brain or liver weight, while the highest dose of MDM1EA decreased body weight and the 2 lowest doses of HMDMA increased body weight. All doses of HMDMA decreased liver weight (expressed as % body weight) when compared with contemporaneous water-treated controls. Taken together, the results of these experiments suggest that structurally related compounds share some stimulus properties with MDMA and may therefore share abuse liability. Furthermore, both MDMA-related compounds produced adverse effects on the developing organism, whereas MDMA did not.

Effects of designer drugs on the chicken embryo and 1-day-old chicken.

The present study was conducted to examine the effects of d-amphetamine and the designer drugs 3,4-methylenedioxymethamphetamine (MDMA), N-methyl-3,4-methylenedioxyphenyl-3-butamine (HMDMA), 3,4-methylenedioxyphenyl-2-butanamine (BDB), 3,4-methylenedioxyphenyl-1-ethanamine (MDM1EA) in the chick embryo and the young chicken. HMDMA and MDM1EA had no effect on motility on day 14 of embryogenesis, while MDMA, BDB, and d-amphetamine decreased embryonic motility at one or more doses. On day 1 posthatch, chickens were challenged with cumulative injections of water or the same drug that they had received in ova. With the exception of MDM1EA, all of the drugs produced effects such as distress vocalization, wing extension, tremor, flat body posture, bursting forward movements, loss of righting reflex, and convulsant-like kicking. Pretreatment with drug in ova resulted in tolerance to certain drug effects and supersensitivity to other drug effects. Furthermore, BDB significantly decreased hatchability, MDM1EA decreased body weight, and HMDMA decreased liver weight. Further studies are needed to determine the mechanism(s) of toxicity in this species.

In vitro aldose reductase inhibitory activity of substituted N-benzenesulfonylglycine derivatives.

A number of N-benzenesulfonylglycines, alanines, sarcosine, and prolines, which contain the minimum pharmacophore moieties necessary for aldose reductase inhibitory activity, were prepared and tested in the rat lens assay. In this assay, the benzenesulfonylglycines are considerably more potent than the corresponding alanine and sarcosine derivatives which, in turn, are more active than the proline analogues. Of the monosubstituted benzenesulfonylglycines, the 2-nitro and 4-amino derivatives were most active with 50% inhibitory concentration (IC50) values of 13 and 16 microM, respectively. The most potent derivatives evaluated were the beta- and alpha-naphthylenesulfonylglycines with IC50 values of 0.4 and 1.3 microM, respectively. The structure-activity data obtained from evaluation of the benzenesulfonylamino acids suggests that the aromatic ring and ring substituents, as well as the sulfonamide group and carboxylate moiety, all contribute to the inhibitory potency through direct interaction with complimentary binding sites present on aldose reductase.

A behavioral comparison of Nexus, cathinone, BDB, and MDA.

The effects of 4-bromo-2,5-dimethoxyphenethylamine (Nexus), 3,4-methylenedioxyamphetamine (MDA), 3,4-methylenedioxyphenyl-2-butanamine (BDB), and cathinone were studied in the newly hatched chicken and compared to the effects of d-amphetamine and three hallucinogens in the same species. Cathinone, a psychomotor stimulant in man (6), produced effects that were qualitatively similar to effects seen after administration of d-amphetamine (i.e., distress vocalization, wing extension, inability to stand, and loss of righting reflex). BDB, a compound with unknown activity in man, and two known hallucinogens, Nexus (5) and MDA (1), produced effects in the chicken that are common to both stimulants and hallucinogens in this species. For example, both MDA and BDB produced abnormal body posture that was identical to that reported after administration of hallucinogens such as lysergic acid diethylamide (LSD) and harmine (11). Nexus, on the other hand, produced rigid penguin-like posture, an effect seen in the chicken after administration of another hallucinogen, mescaline (12). BDB also produced bursting forward movements, an effect commonly observed after LSD and harmine. Our findings suggest that the young chicken can be used as an alternative, nonmammalian, model for predicting classification of new compounds.

Structure-activity relationships of BDB and its monomethyl and dimethyl derivatives.

The purpose of the present study was to evaluate the behavioral effects of 3,4-methylenedioxyphenyl-2-butanamine (BDB), N-methyl BDB (MBDB), and N,N-dimethyl BDB (MMBDB) in the newly hatched chicken. The primary amine, BDB, produced effects that are commonly seen in the chicken after administration of both hallucinogens and psychomotor stimulants (i.e., distress vocalization, tremor, and wing extension). It also produced abnormal body posture and bursting forward locomotion, effects elicited only by hallucinogens. Loss of righting reflex also occurred at the highest (16 mg/kg) dose of BDB, and this effect is typical of d-amphetamine but has not been reported for hallucinogens. The monomethylated derivative of BDB, MBDB, was less potent than BDB, and the N,N-dimethyl analogue of BDB, MMBDB, had no effect on behavior at the doses tested.

In-vitro hydrolysis, permeability, and ocular uptake of prodrugs of N-[4-(benzoylamino)phenylsulfonyl]glycine, a novel aldose reductase inhibitor.

To enhance the ocular uptake of N-[4-(benzoylamino)phenylsulfonyl]glycine (BAPSG), two ester (methyl and isopropyl) prodrugs were synthesized and evaluated for their stability in various buffers (pH 1-9), hydrolysis in rabbit ocular tissues (cornea, conjunctiva, iris-ciliary body, lens, aqueous humor, and vitreous humor), transport across cornea and conjunctiva, and in-vivo uptake following topical administration. Over the pH range of 1-9, the rate constants for degradation ranged from 5.67 to 218.9 x 10(-3) h(-1) for the methyl ester and from 3.14 to 4.45 x 10(-3) h(-1) for the isopropyl ester. At all pH conditions, the isopropyl ester was more stable when compared with the methyl ester. A change in buffer concentration at pH 7.4 did not influence the stability of the prodrugs. The prodrugs were rapidly hydrolysed in the tissue homogenates, with the rate constants for hydrolysis ranging from 1.98 to 7.2x 10(-3) min(-1) for the methyl ester and 3.32 to 6.53 x 10(-3) min(-1) for the isopropyl ester. The in-vitro permeability of the methyl ester was less than the parent drug across cornea and conjunctiva. Isopropyl ester levels were not detectable in the receiver chamber even at the end of the 4-h transport study. Following topical administration of BAPSG and the two prodrugs at a dose of 60 microg/eye, the lowest levels were seen in vitreous humor for parent compound and its methyl ester. In general, the tissue uptake of methyl ester was less than BAPSG. Isopropyl ester levels were below detection limits in all the ocular tissues. Lipophilic ester prodrugs of BAPSG showed good aqueous solution stability in tissue homogenates. However, these prodrugs lacking the free carboxylate anion exhibited reduced in-vitro permeability and in-vivo uptake, suggesting the importance of free carboxylate anion in the delivery of BAPSG.

Rabbit corneal and conjunctival permeability of the novel aldose reductase inhibitors: N-[[4-(benzoylamino)phenyl] sulphonyl]glycines and N-benzoyl-N-phenylglycines.

Corneal and conjunctival permeability has been investigated for novel aldose reductase inhibitors (ARIs) of the N{[4-(benzoylamino)phenyl]sulphonyl}glycine (benzoylaminophenylsulphonylglycine) and N-benzoyl-N-phenylglycine (benzoylphenylglycine) series, compounds developed for prevention of cataract formation in diabetic subjects. Six benzoylaminophenylsulphonylglycines were synthesized with modifications either of the phenyl group or of the glycine structure and three benzoylphenylglycines were synthesized with modification in the phenyl group of the benzoyl moiety. Transport of ARIs in the mucosal to serosal direction was evaluated across rabbit cornea and conjunctiva bathed in glutathione-bicarbonate Ringer's solution maintained at pH 7.4 and 37 degrees C. The permeability coefficients of the novel ARIs across cornea and conjunctiva ranged from 1.87 to 8.95 x 10(-6) cm s(-1) and from 4.6 to 19.15 x 10(-6) cm s(-1), respectively. The ratio of corneal to conjunctival permeability ranged from 0.12 to 0.79. The calculated log partition coefficient (log P) values for the ARIs were in the range 0.84 to 2.78. The log distribution coefficients (log D) were in the range -2.87 to -0.89. There was no apparent relationship between log P or log D and the permeability coefficients of the ARIs for either tissue. Cornea was more resistant to ARI transport than was conjunctiva. Substitution of a phenyl group for hydrogen in the glycine methylene group reduced the permeability coefficient. Permeability coefficients were different for different stereoisomers. Compared with the permeability coefficient of benzoylaminophenylsulphonylglycine, that of 4-fluorobenzoylaminophenylsulphonylglycine was lower in the cornea but similar in the conjunctiva. In both tissues, the permeability coefficient of 2-nitrobenzoylaminophenylsulphonylglycine was less than that of 4-nitrobenzoylaminophenylsulphonylglycine. There was no significant difference between the permeability coefficients of 3-nitro- and 4-nitrobenzoylphenylglycines through either tissue and the permeability coefficients of these compounds were greater than that of the more lipophilic 4-methylbenzoylphenylglycine. The lack of dependence of the permeability coefficients on log P or log D and the different permeabilities of stereoisomers imply the existence of specialized transport processes for the ARIs tested in this study.

Glutamine-supported motility of adult filarial parasites in vitro and the effect of glutamine antimetabolites.

The survival in culture of adult female Brugia pahangi, Acanthocheilonema viteae, and Onchocerca volvulus and adult male Onchocerca gibsoni was assessed by measuring parasite motility. Survival of all species was maximal in a nutritionally complex medium (RPMI-1640). All species survived for up to 48 hr in a simpler medium in which the only energy source was 10 mM glutamine; motility in this medium was dependent upon pH. For the species of Onchocerca, motility was maintained better in the presence of glutamine as the sole energy source than in glucose-only medium. Motility of B. pahangi incubated in 10 mM succinate was equivalent to that seen with 10 mM glutamine, but no other tricarboxylic acid intermediate supported this parasite in vitro. Antimycin A (1 microM) and potassium cyanide (KCN, 100 microM) paralyzed B. pahangi incubated in 10 mM glutamine, an effect antagonized by glucose. KCN at 10 or 100 microM was effective also against Onchocerca gutturosa in glutamine-only medium. Several glutamine antimetabolites reduced motility of B. pahangi by 72 hr. This inhibition was prevented by 2 mM glutamine. However, the inhibition of motility in the species of Onchocerca caused by these compounds was attenuated only partially by glutamine. These data demonstrate that, under certain conditions, filarial nematodes can utilize non-sugar substrates as energy sources. The differential sensitivity seen among these organisms to mitochondrial toxins and glutamine antimetabolites may be related to the extent to which they can use these alternative substrates to generate energy.

Liquid chromatographic determination of the enantiomeric composition of amphetamine prepared from norephedrine and norpseudoephedrine.

The stereochemical course of the synthesis of amphetamine from norephedrine and norpseudoephedrine is investigated using liquid chromatography. The results show that the chiral carbon common to both compounds remains unaffected during the reaction sequence. The presence of individual amphetamine enantiomers in the reaction products is determined by reversed-phase liquid chromatographic separation on an achiral stationary phase (C18) following precolumn derivatization with 2,3,4,6-tetra-O-acetyl-beta-D-glucopyranosyl isothiocyanate (GITC). The GITC derivatization procedure allows for the liquid chromatographic separation of the individual enantiomers of amphetamine, norephedrine, norpseudoephedrine, and the intermediate 1-chloro-1-phenyl-2-aminopropanes.

Liquid chromatographic optimization of reaction conditions for the synthesis of 2,2-dimethyl-N-benzylmalonamide, a new anticonvulsant.

Reversed-phase liquid chromatographic (RP/LC) procedures are used to optimize the reaction conditions for the large scale preparation of 2,2-dimethyl-N-benzylmalonamide, 1. In acute studies using mice and rats, compound 1 is a very effective anticonvulsant agent. Large quantities of 1 (ca. 300 g) are needed for detailed studies of its chronic effects. LC monitoring of the synthesis of intermediate products and 1 result in the optimum use of reagents, increased product yields, and decreased reaction times.

Liquid chromatographic and spectral analysis of the 17-hydroxy anabolic steroids.

The liquid chromatographic properties of various 17-hydroxy anabolic steroids are examined under reversed-phase conditions. These anabolic steroids are now listed as controlled drugs in many states due to their abuse potential in athletics, body building, and other areas. These nonesterified steroids are separated on a C18 stationary phase with a 70% methanol in water mobile phase. In a few cases, two compounds display very similar retention properties. However, dual-wavelength detection at 254 and 280 nm allows for their differentiation. Reversed-phase retention parallels steroid lipophilicity based on hydroxyl and methyl group substituents. Also, those steroids containing a dienone substructure are more polar than steroids containing an enone moiety.